Parkinson's disease (PD) is the second most common neurodegenerative disease in the developed world. Age is the strongest risk factor in PD with incidence rising exponentially after 65 (Caine & Langston. (1983) Lancet, 2, 1457-9; de Lau et al. (2004) Neurology, 63, 1240-4; herein incorporated by reference in their entireties). Because improvements in health care are increasing life expectancy, the number of PD patients is expected to grow dramatically in the coming years, reaching over 2 million in the U.S. by 2025 (Dorsey et al. (2007) Neurology, 68, 384-6; herein incorporated by reference in its entirety). This is expected to have an enormous economic cost, reaching over 50 billion dollars per year by 2025.
Although there are signs of distributed neuropathology in PD (as judged by LB formation) (Braak et al., 2004), the motor symptoms, including bradykinesia, rigidity, and resting tremor, are clearly linked to the degeneration and death of SNc DA neurons (Hornykiewicz. (1966) Pharmacol Rev, 18, 925-64; Riederer & Wuketich. (1976) J Neural Transm, 38, 277-301; herein incorporated by reference in their entireties). The palliative efficacy of L-DOPA—a DA precursor—is testament to the centrality of these neurons in the motor symptoms of PD.
The factors governing the loss of substantia nigra pars compacta (SNc) dopamine (DA) neurons have been the subject of speculation for decades. DA itself has long been viewed as a culprit, as oxidation of cytosolic DA, and its metabolites, is damaging (Greenamyre & Hastings. (2004) Science, 304, 1120-2; Sulzer. (2007) Trends Neurosci, 30, 24450; herein incorporated by reference in their entireties). However, there are reasons to doubt this type of cellular stress alone is responsible for the loss of DA neurons in PD. First, there is considerable regional variability in the vulnerability of DA neurons in PD, with some being devoid of pathological markers (Matzuk & Saper. (1985) Ann Neurol, 18, 552-5; Kish et al. (1988) N Engl J Med, 318, 876-80; Saper et al. (1991) Ann Neurol, 29, 577-84; Ito et al. (1992) Ann Neurol, 32, 543-50; Damier et al. (1999) Brain, 122 (Pt 8), 1437-48; herein incorporated by reference in their entireties). Second, L-DOPA administration, which relieves symptoms by elevating DA levels in PD patients, does not appear to accelerate disease progression, suggesting that DA is not a significant source of reactive oxidative stress, at least in the short term (Fahn. (2005) J Neurol, 252 Suppl 4, IV37-IV42; herein incorporated by reference in its entirety). Calcium entry through L-type channels has been shown to stimulate DA metabolism in SNc DA neurons, pushing cytosolic DA concentrations into a toxic range with L-DOPA loading (Mosharov et al. (2009) Neuron, 62, 218-29; herein incorporated by reference in its entirety). However, the frank death or degeneration of a variety of non-dopaminergic neurons in PD argues that DA itself is not likely to be the principal cell autonomous risk factor in the disease.
Unlike the vast majority of neurons in the brain, adult SNc DA neurons are autonomously active, generating regular, broad action potentials (2-4 Hz) in the absence of synaptic input (Grace & Bunney. (1984) J Neurosci, 4, 2866-76; Nedergaard et al. (1993) J Physiol, 466, 727-47; Guzman et al. (2009) J Neurosci, 29, 11011-9; Chan et al. (2007) Nature, 447, 1081-6; herein incorporated by reference in their entireties). This pace making activity is believed to be important to maintaining ambient DA levels in regions that are innervated by these neurons, particularly the striatum (Romo & Schultz. (1990) J Neurophysiol, 63, 592-606; herein incorporated by reference in its entirety). While most neurons rely exclusively on monovalent cation channels to drive pacemaking, SNc DA neurons also engage L-type ion channels that allow calcium to enter the cytoplasm (Ping & Shepard. (1996) Neuroreport, 7, 809-14; Bonci et al. (1998) J Neurosci, 18, 6693-703; Puopolo et al. (2007) J Neurosci, 27, 645-56; herein incorporated by reference in their entireties), leading to oscillations in intracellular calcium concentrations (Guzman et al. (2009) J Neurosci, 29, 11011-9; Chan et al. (2007) Nature, 447, 1081-6; Wilson & Callaway. (2000) J Neurophysiol, 83, 3084-100; herein incorporated by reference in their entireties). The L-type calcium channels used by SNc DA neurons in pacemaking have a distinctive Cav1.3 pore-forming subunit encoded by Cacna1d. Cav1.3 calcium channels are relatively rare, constituting only about 10% of all the L-type calcium channels found in the brain. Channels with this subunit differ from other L-type calcium channels in that they open at relatively hyperpolarized potentials (Xu & Lipscombe. (2001) J Neurosci, 21, 5944-51; herein incorporated by reference in its entirety), allowing them to contribute to the mechanisms driving the membrane potential to action potential threshold underlying autonomous pacemaking.
The sustained engagement of Cav1.3 calcium channels during pace making comes at an apparent metabolic cost to SNc DA neurons. Because of its involvement in cellular processes ranging from the regulation of enzyme activity to programmed cell death, calcium is under very tight homeostatic control, with a cytosolic set point near 100 nM-10,000 times lower than the concentration of calcium in the extracellular space (Berridge et al. (2000) Nat Rev Mol Cell Biol, 1, 11-21; Rizzuto. (2001) Curr Opin Neurobiol, 11, 306-11; Orrenius et al. (2003) Nat Rev Mol Cell Biol, 4, 552-65; herein incorporated by reference in their entireties). Calcium entering neurons is rapidly sequestered or pumped back across the steep plasma membrane concentration gradient; this process requires energy stored in adenosine triphosphate (ATP) or in ion gradients that are maintained with ATP-dependent pumps, like the Na-K ATPase. In most neurons, calcium channel opening is a rare event, occurring primarily during very brief action potentials. This makes the metabolic cost to the cell readily manageable. But in SNc DA neurons, where Cav1.3 calcium channels are open much of the time, the magnitude and the spatial extent of calcium influx are much larger. Transgenic mice that express a mitochondrially targeted redox-sensitive variant of green fluorescent protein (mito-roGFP) under control of the tyrosine hydroxylase promoter have revealed that indeed mitochodria in SNc DA neurons have a high basal oxidant stress that is a direct consequence of opening of L-type calcium channels. Furthermore, calcium entry (and presumably the concomitant oxidant stress) increases the vulnerability of SNc DA neurons to toxins (MPTP, 6-hydroxydopamine (6-OHDA), rotenone) used to create animal models of PD.
These results indicate that calcium entry during pace making elevates mitochondrial oxidant stress in SNc DA neurons and increases their vulnerability to toxins and genetic mutations with metabolic consequences. This oxidant stress also should increase the rate of cellular aging and death. This physiological model of vulnerability in PD and other aging related neurodegenerative diseases extends beyond SNc DA neurons to ex-plain the loss of many of the other neuronal populations lost in the disease, including neurons of the locus ceruleus, raphe nuclei, hypothalamus, and penduculopontine nucleus.
Epidemiological studies have shown that use of dihydropyridines that cross the blood-brain barrier are associated with a significant reduction in the risk of developing Parkinson's disease. The utility of these compounds is limited by their cardiovascular side-effects, which are attributable to their antagonism of Cav1.2 channels in heart and vascular smooth muscle. Due to their affinity for Cav1.2 channels, at higher doses, dihydropyridines induce hypotension and cardiac failure. This severely limits their utility as a neuroprotective agent. What is needed are pharmacological agents that preferentially antagonize calcium channels with a Cav1.3 pore, the type of channels responsible for neurodegeneration in PD.